Of the species examined, the rat, a corticosterone-producing animal, was found to have the highest specific activity of AHH in the adrenal gland, whereas a low or no activity was found in three adult cortisol-producing animals, the guinea pig, dog, and man. In the rat adrenal, AHH activity was localized predominantly in the zona fasciculata, and was not inducible by 3-methyl cholanthrene or TCDD (2,3,7,8-tetrachlorodibenzo-p-dioxin) treatment, but only by ACTH administration. Following ACTH treatment of rats, the increase in AHH activity was concurrent with a similar increase in the concentration of an apoprotein of cytochrome P-450 in the molecular weight region of 54,000. The latter apoprotein of P-450 was absent in the adult but not the fetal human adrenal and its absence correlated with the lack of AHH activity, but not steroid hydroxylase activity. The latter finding suggests that the P-450 apoprotein of 54,000 molecular weight may be specific for AHH activity.